IND-enabling Studies of ZB716, an Orally Bioavailable SERD Project Summary Most patients with advanced metastatic breast cancer eventually develop resistance to tamoxifen or aromatase inhibitor (AI) treatment where the recurrent and/or progressive disease retains the expression of ER?. The standard treatment of breast cancer progressing after tamoxifen or AI therapy is fulvestrant which is the only FDA approved selective estrogen receptor degrader (SERD) as a second-line endocrine regimen. Due to its extremely poor oral bioavailability, fulvestrant was administered as a 250 mg/month by intramuscular injection, which was approved in 2002. It takes 3-4 month to reach the steady state serum concentration of fulvestrant at 15 ng/mL in patients. Subsequent clinical trials using 500mg/month with an additional loading dose on day 14 demonstrated significant clinical improvement, leading to the 2011 FDA approval of fulvestrant as a 500 mg injection regimen. However, even at this dosage, the peak blood concentration of fulvestrant remains below a modest 25 ng/mL, and the time to steady-state drug concentration remains about 30 days. These shortcomings of fulvestrant may account for the limited clinical efficacy low patient response rate. Thus, a potent, orally bioavailable SERD has potential for significantly higher receptor knockdown and for more durable clinical benefits than fulvestrant. To date, only two nonsteroidal oral SERDs, GDC-0810 (Genentech) and AZD9496 (AstraZeneca) are being tested in clinical trials. These oral SERDs have very different molecular structures from fulvestrant, are less potent than fulvestrant in preclinical studies, and are at least several years away from being proven clinically safe and efficacious. On the other hand, few reports have described attempts to make orally bioavailable steroidal SERDs and none has progressed to clinical studies. Indeed, these attempts focused on modifications made primarily to the long alkyl chain to increase polarity and solubility but failed to address the main problem that is responsible for the poor bioavailability of fulvestrant, that is, fulvestrant undergoes rapid and extensive O-glucuronidation and O-sulfation to form polar metabolites that are inactive and water soluble. Zenopharm has developed ZB716, a patented steroidal oral SERD that can effectively enhance systemic bioavailability while bypassing first-pass metabolism (glucuronidation and sulfation) of fulvestrant. Preclinical studies confirmed that this chemical modification can retain sufficiently high binding affinity of the steroidal moiety of fulvestrant while minimizing glucuronidation and sulfation. We found that ZB716 binds to ER with high affinity and exerts its antiestrogenic effect on ER-expressing breast cancer cells. In both tamoxifen naive and tamoxifen resistant breast cancer cells, ZB716 potently inhibits cell proliferation and effectively degrades the hormone receptor in a dose- dependent manner. Moreover, ZB716 is shown to have far superior oral bioavailability in mice compared to fulvestrant. Therefore ZB716 is a viable oral SERD, which can not only overcome the disadvantages associated with injection depot, but more importantly can improve therapeutic efficacy and achieve more durable treatment outcome than the current SERD regimen. To move ZB716 towards clinical trials we propose to conduct IND-enabling studies that will initiate CMC (chemistry, manufacturing, and control) work and investigate the in vivo efficacy of oral ZB716 in two xenograft models. The dose-dependent efficacy studies are the next key steps to determine if the oral bioavailability of ZB716 can be translated to in vivo efficacy. Optimization of synthetic procedure for use in scale-up preparation of GLP and GMP grade ZB716 is necessary to custom-manufacture the API.